Computational analyses of curcuminoid analogs against kinase domain of HER2.

BACKGROUND: Human epidermal growth factor receptor 2 (HER2) has an important role in cancer aggressiveness and poor prognosis. HER2 has been used as a drug target for cancers. In particular, to effectively treat HER2-positive cancer, small molecule inhibitors were developed to target HER2 kinase. Knowing that curcumin has been used as food to inhibit cancer activity, this study evaluated the efficacy of natural curcumins and curcumin analogs as HER2 inhibitors using in vitro and in silico studies. The curcumin analogs considered in this study composed of 4 groups classified by their core structure, ß-diketone, monoketone, pyrazole, and isoxazole. RESULTS: In the present study, both computational and experimental studies were performed. The specificity of curcumin analogs selected from the docked results was examined against human breast cancer cell lines. The screened curcumin compounds were then subjected to molecular dynamics simulation study. By modifying curcumin analogs, we found that protein-ligand affinity increases. The benzene ring with a hydroxyl group could enhance affinity by forming hydrophobic interactions and the hydrogen bond with the hydrophobic pocket. Hydroxyl, carbonyl or methoxy group also formed hydrogen bonds with residues in the adenine pocket and sugar pocket of HER2-TK. These modifications could suggest the new drug design for potentially effective HER2-TK inhibitors. Two outstanding compounds, bisdemethylcurcumin (AS-KTC006) and 3,5-bis((E)-3,4-dimethoxystyryl)isoxazole (AS-KTC021 ),were well oriented in the binding pocket almost in the simulation time, 30 ns. This evidence confirmed the results of cell-based assays and the docking studies. They possessed more distinguished interactions than known HER2-TK inhibitors, considering them as a promising drug in the near future. CONCLUSIONS: The series of curcumin compounds were screened using a computational molecular docking and followed by human breast cancer cell lines assay. Both AS-KTC006 and AS-KTC021 could inhibit breast cancer cell lines though inhibiting of HER2-TK. The intermolecular interactions were confirmed by molecular dynamics simulation studies. This information would explore more understanding of curcuminoid structures and HER2-TK.

Discovery of 7-aryl-substituted (1,5-naphthyridin-4-yl)ureas as aurora kinase inhibitors.

As part of our research projects to identify new chemical entities of biological interest, we developed a synthetic approach and the biological evaluation of (7-aryl-1,5-naphthyridin-4-yl)ureas as a novel class of Aurora kinase inhibitors for the treatment of malignant diseases based on pathological cell proliferation. 1,5-Naphthyridine derivatives showed excellent inhibitory activities toward Aurora kinases A and B, and the most active compound, 1-cyclopropyl-3-[7-(1-methyl-1H-pyrazol-4-yl)-1,5-naphthyridin-4-yl]urea (49), displayed IC50 values of 13 and 107 nM against Aurora kinases A and B, respectively. In addition, the selectivity toward a panel of seven cancer-related protein kinases was highlighted. In vitro ADME properties were also determined in order to rationalize the difficulties in correlating antiproliferative activity with Aurora kinase inhibition. Finally, the good safety profile of these compounds imparts promising potential for their further development as anticancer agents.

Monocarbonyl curcumin analogues: heterocyclic pleiotropic kinase inhibitors that mediate anticancer properties.

Curcumin is a biologically active component of curry powder. A structurally related class of mimetics possesses similar anti-inflammatory and anticancer properties. Mechanism has been examined by exploring kinase inhibition trends. In a screen of 50 kinases relevant to many forms of cancer, one member of the series (4, EF31) showed ≥85% inhibition for 10 of the enzymes at 5 µM, while 22 of the proteins were blocked at ≥40%. IC50 values for an expanded set of curcumin analogues established a rank order of potencies, and analyses of IKKß and AKT2 enzyme kinetics for 4 revealed a mixed inhibition model, ATP competition dominating. Our curcumin mimetics are generally selective for Ser/Thr kinases. Both selectivity and potency trends are compatible with protein sequence comparisons, while modeled kinase binding site geometries deliver a reasonable correlation with mixed inhibition. Overall, these analogues are shown to be pleiotropic inhibitors that operate at multiple points along cell signaling pathways.

Sphingosine and FTY720 are potent inhibitors of the transient receptor potential melastatin 7 (TRPM7) channels.

BACKGROUND AND PURPOSE: Transient receptor potential melastatin 7 (TRPM7) is a unique channel kinase which is crucial for various physiological functions. However, the mechanism by which TRPM7 is gated and modulated is not fully understood. To better understand how modulation of TRPM7 may impact biological processes, we investigated if TRPM7 can be regulated by the phospholipids sphingosine (SPH) and sphingosine-1-phosphate (S1P), two potent bioactive sphingolipids that mediate a variety of physiological functions. Moreover, we also tested the effects of the structural analogues of SPH, N,N-dimethyl-D-erythro-sphingosine (DMS), ceramides and FTY720 on TRPM7. EXPERIMENTAL APPROACH: HEK293 cells stably expressing TRPM7 were used for whole-cell, single-channel and macropatch current recordings. Cardiac fibroblasts were used for native TRPM7 current recording. KEY RESULTS: SPH potently inhibited TRPM7 in a concentration-dependent manner, whereas S1P and other ceramides did not produce noticeable effects. DMS also markedly inhibited TRPM7. Moreover, FTY720, an immunosuppressant and the first oral drug for treatment of multiple sclerosis, inhibited TRPM7 with a similar potency to that of SPH. In contrast, FTY720-P has no effect on TRPM7. It appears that SPH and FTY720 inhibit TRPM7 by reducing channel open probability. Furthermore, endogenous TRPM7 in cardiac fibroblasts was markedly inhibited by SPH, DMS and FTY720. CONCLUSIONS AND IMPLICATIONS: This is the first study demonstrating that SPH and FTY720 are potent inhibitors of TRPM7. Our results not only provide a new modulation mechanism of TRPM7, but also suggest that TRPM7 may serve as a direct target of SPH and FTY720, thereby mediating S1P-independent physiological/pathological functions of SPH and FTY720.

Blockade of STAT3 activation by sorafenib derivatives through enhancing SHP-1 phosphatase activity.

Previously, we demonstrated that the multiple kinase inhibitor sorafenib mediates the repression of phospho-STAT3 in hepatocellular carcinoma cells. In this study, we used this kinase-independent mechanism as a molecular basis to use sorafenib as scaffold to develop a novel class of SHP-1-activating agents. The proof of principle of this premise was provided by SC-1, which on replacement of N-methylpicolinamide by a phenylcyano group showed abolished kinase activity while retaining phospho-STAT3 repressive activity. Structural optimization of SC-1 led to compound 6, which repressed phospho-STAT3 through SHP-1 activation and inhibited PLC5 cell proliferation at sub-micromolar potency. In light of the pivotal role of phospho-STAT3 in promoting tumorigenesis and drug resistance, this novel SHP-1-activating agent may have therapeutic relevance in cancer therapy.

Docking, synthesis and pharmacological activity of novel urea-derivatives designed as p38 MAPK inhibitors.

p38 mitogen-activated protein kinase (p38 MAPK) is an important signal transducing enzyme involved in many cellular regulations, including signaling pathways, pain and inflammation. Several p38 MAPK inhibitors have been developed as drug candidates to treatment of autoimmune disorders, such as rheumatoid arthritis. In this paper we reported the docking, synthesis and pharmacological activity of novel urea-derivatives (4a-e) designed as p38 MAPK inhibitors. These derivatives presented good theoretical affinity to the target p38 MAPK, standing out compound 4e (LASSBio-998), which showed a better score value compared to the prototype GK-00687. This compound was able to reduce in vitro TNF-α production and was orally active in a hypernociceptive murine model sensible to p38 MAPK inhibitors. Otherwise, compound 4e presented a dose-dependent analgesic effect in a model of antigen (mBSA)-induced arthritis and anti-inflammatory profile in carrageenan induced paw edema, indicating its potential as a new antiarthritis prototype.

Fluorescence polarization assay for inhibitors of the kinase domain of receptor interacting protein 1.

Necrotic cell death is prevalent in many different pathological disease states and in traumatic injury. Necroptosis is a form of necrosis that stems from specific signaling pathways, with the key regulator being receptor interacting protein 1 (RIP1), a serine/threonine kinase. Specific inhibitors of RIP1, termed necrostatins, are potent inhibitors of necroptosis. Necrostatins are structurally distinct from one another yet still possess the ability to inhibit RIP1 kinase activity. To further understand the differences in the binding of the various necrostatins to RIP1 and to develop a robust high-throughput screening (HTS) assay, which can be used to identify new classes of RIP1 inhibitors, we synthesized fluorescein derivatives of Necrostatin-1 (Nec-1) and Nec-3. These compounds were used to establish a fluorescence polarization (FP) assay to directly measure the binding of necrostatins to RIP1 kinase. The fluorescein-labeled compounds are well suited for HTS because the assays have a dimethyl sulfoxide (DMSO) tolerance up to 5% and Z' scores of 0.62 (fluorescein-Nec-1) and 0.57 (fluorescein-Nec-3). In addition, results obtained from the FP assays and ligand docking studies provide insights into the putative binding sites of Nec-1, Nec-3, and Nec-4.

Chemistry and biology of fascaplysin, a potent marine-derived CDK-4 inhibitor.

Marine natural products offer an abundant source of pharmacologically active agents with great diversity and complexity, and the potential to produce valuable therapeutic entities. Indole alkaloids is one of the important class of marine-derived secondary metabolites, with wide occurrence amongst variety of marine sources such as sponges, tunicates, algae, worms and microorganisms and have been extensively studied for their biological activities. Among this chemical family, a sponge-derived bis-indole alkaloid fascaplysin (1) exhibited broad range of bioactivities including antibacterial, antifungal, antiviral, anti-HIV-1-RTase, p56 tyrosine kinase inhibition, antimalarial, anti-angiogenic, antiproliferative activity against numerous cancer cell lines, specific inhibition of cyclin-dependent kinase-4 (IC(50) 350 nM) and action as a DNA intercalator. In the present review, the chemical diversity of natural as well as synthetic analogues of fascaplysin has been reviewed with a detailed account on synthetic reports and pharmacological studies. Our analysis of the structure-activity relationships of this family of compounds highlights the existence of various potential leads for the development of novel anticancer agents.

Meridianins: marine-derived potent kinase inhibitors.

Marine invertebrates are a rich source of novel, bioactive secondary metabolites and have attracted a great deal of attention from scientists in the fields of chemistry, pharmacology, ecology, and molecular biology. This profilic natural source has produced several antitumor secondary metabolites and amongst these, indole alkaloids are of wide occurrence. Meridianins A-G (1-7) are indole alkaloids isolated from tunicate Aplidium meridianum and are known to inhibit variety of protein kinases associated with cancer and neurodegenerative diseases. These compounds also exhibited promising antiproliferative activity in several cancer cell lines. Amongst natural meridianins, meridianin E (5) showed potent and selective inhibition of CDK-1 and CDK-5. Several synthetic meridianin analogs exhibited potent and selective inhibition of glycogen synthase-3 (GSK-3) and dual-specificity tyrosine-phosphorylation regulated kinase 1A (Dyrk-1A) which are known to be implicated in progression of Alzheimer's disease. The present review provides the critical account of isolation, medicinal chemistry and pharmacology of meridianins. Our analysis of the structure-activity relationships of this family of compounds highlights the existence of various potential leads for the development of novel anticancer and anti-Alzheimer's agents.

Synthesis and inhibitory activity of thymidine analogues targeting Mycobacterium tuberculosis thymidine monophosphate kinase.

We report on Mycobacterium tuberculosis thymidine monophosphate kinase (TMPKmt) inhibitory activities of a series of new 3'- and 5'-modified thymidine analogues including α- and ß-derivatives. In addition, several analogues were synthesized in which the 4-oxygen was replaced by a more lipophilic sulfur atom to probe the influence of this modification on TMPKmt inhibitory activity. Several compounds showed an inhibitory potency in the low micromolar range, with the 5'-arylthiourea 4-thio-α-thymidine analogue being the most active one (K(i)=0.17 µM). This compound was capable of inhibiting mycobacteria growth at a concentration of 25 µg/mL.

A practical approach to new (5Z) 2-alkylthio-5-arylmethylene-1-methyl-1,5-dihydro-4H-imidazol-4-one derivatives.

A practical protocol for the preparation of (5Z)-2-alkylthio-5-arylmethylene-1-methyl-1,5-dihydro-4H-imidazol-4-one derivatives is reported. The new compounds were obtained in good yield and stereoselectivity in two steps, namely a solvent-free Knoevenagel condensation under microwave irradiation, followed by an S-alkylation reaction with various halogenoalkanes.

Anti-angiogenic effects of purine inhibitors of cyclin dependent kinases.

Small molecular inhibitors of Cyclin dependent kinases (Cdks) are currently being developed as anticancer therapeutics due to their antiproliferative properties. The purine Cdk specific inhibitor (R)-roscovitine (seliciclib, CYC202) represents one of the most promising of these compounds. It is currently evaluated in clinical trials concerning cancer therapy. Recently, we have shown that roscovitine exerts potent antiangiogenic effects and elucidated Cdk5 as a new player in angiogenesis. These findings introduce Cdk5 as novel target for antiangiogenic therapy, and Cdk5 inhibitors as an attractive therapeutic approach. Here, we present the antiangiogenic profile of 15 derivatives of roscovitine in vitro and in vivo and provide structure activity relationships of the roscovitine analogs. The (S)-isomer LGR561 and the respective (R)- and (S)-isomers LGR848 and LGR849 strongly inhibited proliferation and cell cycle progression, induced cell death, and reduced migration of endothelial cells in vitro. In comparison to roscovitine, these compounds showed an increased potency to inhibit Cdk2, Cdk5, Cdk7, and Cdk9. By analyzing the effects of LGR561, LGR848, and LGR849 on endothelial cell tube formation, mouse aortic ring sprouting, angiogenesis in the chick chorioallantoic membrane, and neovessel formation in the mouse cornea, we elucidate the two (S)-isomers LGR561 and LGR849 as highly potent inhibitors of angiogenesis. This study provides first information on how to modify roscovitine to develop Cdk inhibitors with increased antiangiogenic activity and suggests the application of existing and the development of new Cdk inhibitors to inhibit both, cancer cell proliferation and angiogenesis.

Direct arylation of simple azoles catalyzed by 1,10-phenanthroline containing palladium complexes: an investigation of C4 arylation of azoles and the synthesis of triarylated azoles by sequential arylation.

Direct triarylation and sequential triarylation reactions of simple azoles catalyzed by [Pd(phen)(2)]PF(6) are described. Simple azoles, such as N-methylimidazole, thiazole, and oxazole, were observed to undergo triaryaltion reactions even at their C4 positions when treated with aryl iodides in the presence of [Pd(phen)(2)]PF(6) as a catalyst and a stoichiometric amount of Cs(2)CO(3) in DMA at 150 °C. Using excess amounts of azoles, selective C5 monoarylation was achieved by using the same catalytic system. Subsequent efforts demonstrated that C5 arylated azoles undergo exclusive C2 arylation using [Pd(phen)(2)]PF(6) as the catalyst with galvinoxyl as an additive. Finally, unprecedented C4 arylation reactions of 2,5-diaryl-azoles occur by using the new catalytic system to give the corresponding triarylated products in good to excellent yields. The results of mechanistic studies suggest that the C2 arylation process takes place by way of an electrophilic aromatic substitution (S(E)Ar) palladation pathway, while arylation reactions at the C4 position occur via a S(E)Ar palladation and/or radical mechanism. Finally, a concise, three-step synthesis of the Tie-2 Tyrosine Kinase Inhibitor has been executed starting with commercially available N-methylimidazole by a route that employs the new sequential arylation process.

Selectivity filters to edit out deleterious side effects in kinase inhibitors.

As the molecular etiology of cancer unravels, revealing the heterogeneous nature of the malignancy, multi-target drug treatments are more frequently advocated. Such therapeutic avenues often target kinases, the basic signal transducers in the cell. Because kinases share common evolutionary backgrounds, they also share many structural attributes, making it difficult for molecular targeted therapy to distinguish between paralogs. Thus, kinase inhibitors (KIs) tend to have undesired cross-reactivities, resulting in potentially lethal side effects. The health risks are obviously higher in these multi-pronged treatments when contrasted with the effects of more selective therapeutic agents. Using a nonconserved physicochemical biomarker, we present a rationally designed molecular filter that enables the control of specificity and the development of adjuvant drugs to edit out the side effects of the primary therapeutic agent. These editors work by overlapping therapeutically with the primary drug in cancer cells, while interfering with toxicity-related signaling pathways recruited by the primary drug in off-target cells. We then examine the possible application of these filtering methods to specifically target kinases when they present idiosyncratic cancer-related mutations. Such applications open the door to engineer personalized drugs tailored to the genetic makeup of the patient. These various methods of enhancing efficacy and safety show some degree of modularity, allowing drug designers to utilize multiple techniques and various drug combinations to create the safest and most powerful treatment for any given therapeutic scenario.

Harmine is an ATP-competitive inhibitor for dual-specificity tyrosine phosphorylation-regulated kinase 1A (Dyrk1A).

Harmine is a ß-carboline alkaloid. The compound is a potent inhibitor of dual-specificity tyrosine phosphorylation-regulated kinase 1A (Dyrk1A), a kinase implicated in Down syndrome. In this study, we show that harmine functions as an ATP-competitive inhibitor against Dyrk1A. Our conclusion is supported by kinetic analysis of harmine inhibition as well as by the characterization of a Dyrk1A mutation conferring significant resistance to harmine. The mutation, V306A, is located next to the highly conserved D307 residue in kinases known to coordinate the phosphate groups of ATP through a Mg²+ ion. The V306A mutation offers harmine resistance by differentially altering Dyrk1A affinity for harmine and ATP. The V306A mutation causes no apparent alteration to Dyrk1A activity except for the reduction in ATP affinity. This deficiency could be fully compensated by supplying ATP with a concentration in the physiological range. Our results reveal that harmine inhibits Dyrk1A activity by interacting with residues in the ATP-binding pocket and displacing ATP. Our results also suggest that harmine will be a good lead compound for further designing of selective ATP-competitive Dyrk1A inhibitors through exploration of the ATP-binding pocket of Dyrk1A.

Evaluation of radioiodinated quinazoline derivative as a new ligand for EGF receptor tyrosine kinase activity using SPECT.

OBJECTIVE: A radioiodinated analog of PD153035 (m-IPQ) was evaluated as a potential epidermal growth factor receptor tyrosine kinase (EGFR-TK) activity imaging ligand for SPECT. METHODS: The 50% inhibition concentration (IC50) value of m-IPQ for EGFR-TK phosphorylation inhibition was evaluated and compared to various EGFR-TK inhibitors. [(125)I]m-IPQ was synthesized by iododestannylation reaction. Biodistribution study of [(125)I]m-IPQ was conducted in normal mice and tumor-bearing mice. The selectivity and binding characteristics (B(max) and K(d)) were analyzed. RESULTS: The quinazoline derivative m-IPQ was found to have high inhibitory potency (IC50: 50.5 ± 3.5 nM) and selectivity toward EGFR-TK. In vivo biodistribution studies of [(125)I]m-IPQ demonstrated its rapid clearance and low retention in normal tissue. On the other hand, high tumor uptake was observed. However, the increase in [(125)I]m-IPQ uptake in the stomach as a deiodination parameter was found. Thus, [(125)I]m-IPQ showed low in vivo stability. The selectivity toward EGFR-TK of m-IPQ was confirmed by the pretreatment experiment with EGFR-TK specific inhibitors, PD153035, Genistein. [(125)I]m-IPQ bound to single population of binding sites with high affinity and kinetic parameter. In addition, [(125)I]m-IPQ was bound to EGFR-TK according to the amount of EGFR-TK expression in the tumor. CONCLUSIONS: [(125)I]m-IPQ showed a relatively high tumor accumulation with selective EGFR-TK binding. Moreover, the tumor uptake of [(125)I]m-IPQ might be reflected in the amount of EGFR-TK expression in the tumor. These good characteristics of [(125)I]m-IPQ suggested that a ¹²³I-labeled counterpart, [¹²³I]m-IPQ, would have great potential for EGFR-TK imaging with SPECT. However, the in vivo stability of this compound needs to improve.

Identification of potent ITK inhibitors through focused compound library design including structural information.

A series of novel compound libraries inhibiting interleukin-2 inducible T cell kinase (ITK) were designed, synthesized and evaluated. In the first design cycle two library scaffolds were identified showing low micromolar inhibition of ITK. Further iterative design cycles including crystal structure information of ITK and structurally related kinases led to the identification of indolylindazole and indolylpyrazolopyridine compounds with low nanomolar ITK inhibition.

Non-ATP competitive protein kinase inhibitors.

Protein kinases represent an attractive target in oncology drug discovery. Most of kinase inhibitors are ATP-competitive and are called type I inhibitors. The ATP-binding pocket is highly conserved among members of the kinase family and it is difficult to find selective agents. Moreover, the ATP-competitive inhibitors must compete with high intracellular ATP levels leading to a discrepancy between IC50s measured by biochemical versus cellular assays. The non-ATP competitive inhibitors, called type II and type III inhibitors, offer the possibility to overcome these problems. These inhibitors act by inducing a conformational shift in the target enzyme such that the kinase is no longer able to function. In the DFG-out form, the phenylalanine side chain moves to a new position. This movement creates a hydrophobic pocket available for occupation by the inhibitor. Some common features are present in these inhibitors. They contain a heterocyclic system that forms one or two hydrogen bonds with the kinase hinge residue. They also contain a hydrophobic moiety that occupies the pocket formed by the shift of phenylalanine from the DFG motif. Moreover, all the inhibitors bear a hydrogen bond donor-acceptor pair, usually urea or amide, that links the hinge-binding portion to the hydrophobic moiety and interacts with the allosteric site. Examples of non ATP-competitive inhibitors are available for various kinases. In this review small molecules capable of inducing the DFG-out conformation are reported, especially focusing on structural feature, SAR and biological properties.

Preparation of novel alkylated arginine derivatives suitable for click-cycloaddition chemistry and their incorporation into pseudosubstrate- and bisubstrate-based kinase inhibitors.

Efficient strategies for the introduction of arginine residues featuring acetylene or azide moieties in their side chains are described. The substituents are introduced in a way that maintains the basicity of the guanidine moiety. The methodology can be used e.g. for non-invasive labeling of arginine-containing peptides. Its applicability is demonstrated by the introduction of 'click' handles into a Protein Kinase C (PKC) pseudosubstrate peptide, and the subsequent preparation and evaluation of a novel bisubstrate-based inhibitor based on such a peptide.